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Patients with chronic kidney disease (CKD) often exhibit increased level of oxidative stress that contribute to the deterioration of renal function and uremic complications. White adipose tissue (WAT) has been recognized as a major site of production of radical oxygen species (ROS) in the context of metabolic diseases. This study was designed to decipher whether the protein bound uremic toxin p-cresyl-sulfate (p-CS) could contribute to ROS production in WAT and promote oxidative stress. Mouse 3T3-L1 adipocytes were incubated for 2 h in culture medium containing 212 μM p-CS, a concentration chosen to mimic levels encountered in end stage renal disease patients or KCl as a control and intracellular ROS production was measured using the fluorescent probe 5-6-carboxy-2′,7′-dichlorodihydrofluorescein diacetate. Oxidative insult was estimated by the measurement of malondialdehyde (MDA) content and glutathione content. The effects of probenecid (1 mM) a potent inhibitor of organic anion transporter, apocynin (1 mM) an inhibitor of NADPH oxidase or common antioxidants such as α-tocopherol (2.5 μM), ascorbate (200 μM), and N-acetylcysteine (500 μM) were further evaluated. p-CS triggered a striking increase in ROS production (+228%, p < 0.01), in MDA content (+214%, p < 0.005) and a decrease in glutathione (−47%, P < 0.01). Pre-treatment of cells with probenecid, apocynin or antioxidants prevented the p-CS induced ROS production and oxidative insults. These results suggest that in uremic state, the intracellular accumulation of p-CS in adipose cells could contribute, through an activation of NADPH oxidase, to the redox imbalance often reported in CKD patients. •p-cresyl-sulfate (p-CS) is a major uremic toxin that accumulates in patients with renal failure. • p-CS promotes ROS production by adipose cells through activation of NADPH oxidase and a decrease in intracellular glutathione pool. • p-CS acts intracellularly to increase ROS production. • Common antioxidants can prevent p-CS induced oxidative stress. [ABSTRACT FROM AUTHOR]

p -Cresyl glucuronide (p-CG) is a by-product of tyrosine metabolism that accumulates in patients with end-stage renal disease. p-CG binding to human serum albumin in physiological conditions (37 °C, pH 7.40) was studied by ultrafiltration (MWCO 10 kDa) and data were analyzed assuming one binding site. The estimated value of the association constant was 2.77 × 10 3  M −1 and a maximal stoichiometry of 3.80 mol per mole. At a concentration relevant for end-stage renal patients, p-CG was 23% bound to albumin. Competition experiments, using fluorescent probes, demonstrated that p-CG did not bind to Sudlow's site I or site II. The p-CG did not interfere with the binding of p-cresyl-sulfate or indoxyl sulfate to serum albumin. [ABSTRACT FROM AUTHOR]

Little is known about potential differences in binding characteristics of protein-bound uremic toxins (PBUTs) in patients with chronic kidney disease (CKD) versus healthy controls. The question arises whether eventual differences are attributed to (i) the elevated levels of competing uremic toxins, and/or (ii) post-translational modifications of albumin. We evaluated the binding characteristics of hippuric acid (HA), indole-3-acetic acid (IAA), indoxyl sulfate (IS), and p -cresylsulfate ( p CS) by deriving a binding curve in three distinct conditions: (i) serum from healthy controls (healthy serum), (ii) blank serum from hemodialysis patients (blank HD serum; i.e. cleared from uremic toxins), and (iii) non-treated serum from HD patients (HD serum). Additionally, the mutual binding competition of these uremic toxins was studied in blank HD in pairs. In both experiments, equilibrium dialysis (37 °C, 5 h) was used to separate the free and bound fractions of each PBUT. Free and total PBUT concentrations were quantified by an ultra-high performance liquid chromatography method with tandem mass spectrometer detection and the percentage protein binding (%PB) of each PBUT was calculated. For all four compounds, the binding capacity of healthy serum was higher than blank HD serum, which was comparable to non-treated HD serum, except for HA. The competition experiments revealed that at high uremic concentrations, mutual competition was observed for the strongly bound PBUTs IS and p CS. The %PB of the weakly bound HA and IAA was lower (trend) only for the addition to blank HD serum containing the strongly bound IS or pCS. There is an intrinsic impact on protein binding in uremia, revealing a lower binding capacity, as compared to healthy controls. Competitive binding is only relevant for the strongly bound PBUTs at high uremic concentrations. In addition, at least part of the effect on binding capacity can be attributed to post-translational modifications of albumin. [ABSTRACT FROM AUTHOR]

Uremic toxins are compounds normally excreted in urine that accumulate in patients with chronic kidney disease as a result of decreased renal clearance. Phenylacetic acid (PAA) has been identified as a new protein bound uremic toxin. The purpose of this study was to investigate in vitro the interaction between PAA and human serum albumin (HSA) at physiological and pathological concentrations. We used ultrafiltration to show that there is a single high-affinity binding site for PAA on HSA, with a binding constant on the order of 3.4 × 10 4 M −1 and a maximal stoichiometry of 1.61 mol per mole. The PAA, at the concentration reported in end-stage renal patients, was 26% bound to albumin. Fluorescent probe competition experiments demonstrated that PAA did not bind to Sudlow's site I (in subdomain IIA) and only weakly bind to Sudlow's site II (in subdomain IIIA). The PAA showed no competition with other protein-bound uremic toxins such as p -cresyl-sulfate or indoxyl sulfate for binding to serum albumin. Our results provide evidence that human serum albumin can act as carrier protein for phenylacetic acid. [ABSTRACT FROM AUTHOR]

White adipose tissue secretes a large variety of compounds named adipokines amongst which, leptin exhibits pleiotropic metabolic actions. Leptin is an anorexigenic hormone, secreted in proportion of fat mass, with additional effects on the regulation of inflammation, cardiovascular system, immunity, hematopoiesis and bone metabolism. Chronic kidney disease (CKD) is characterized by an increase of plasma leptin concentration that may be explained by a lack of renal clearance. Hyperleptinemia plays a key role in the pathogenesis of complications associated with CKD such as cachexia, protein energy wasting, chronic inflammation, insulin resistance, cardiovascular damages and bone complications. Leptin is also involved in the progression of renal disease through its pro-fibrotic and pro-hypertensive actions. Most of the adverse effects of leptin have been documented both experimentally and clinically. Leptin may therefore be considered as an uremic toxin in CKD. The aim of this review is to summarize the pathophysiological and clinical role of leptin in in vitro studies, experimental models, as well as in patients suffering from CKD. [ABSTRACT FROM AUTHOR]